Effect of Hypericum thymbrifolium BOISS. ET NOE, Hypericum scabrum L. and Eryngium creticum LAM. plant extracts on Leishmania major, Leishmania tropica and Leishmania infantum/donovani strains and their cytotoxic potential.

Leishmaniasis causes significant morbidity and mortality worldwide. In our country, there has been a significant increase in the number of cases of leishmaniasis in the last decade. In our study, the effects of Hypericum thymbrifolium, Hypericum scabrum and Eryngium creticum plant extracts were tested on Leishmania major, Leishmania tropica and Leishmania infantum/donovani, which were clinically resistant by not responding to Glucantime® therapy. Cytotoxicity of these extracts were evaluated by XTT method in the human fibroblast cell line. Possible active ingredients were detected by GC-MS analysis from plant extracts. Glucantime® resistance was detected at concentrations of 50 µg/mL and lower in 4 of the 7 strains tested. No living leishmania parasites were found in leishmania strains treated with plant extracts at concentrations of 100 µg/mL or higher. The concentrations of plant extracts included in the study on the WI-38 human fibroblast cell line were not cytotoxic. According to the GC-MS analysis, several active substances with biological activities and anti-parasitic effects, such as Thiophene, Germacrene-D, trans-Geranylgeraniol, Pyridine, and Maleimides, were identified. Based on the findings of the study, it is believed that these identified active substances when supported by in-vivo studies, will pave the way for future research and have the potential to be developed as anti-leishmania drugs.

In vitro and in vivo anti-parasitic activity of curcumin nanoemulsion on Leishmania major (MRHO/IR/75/ER).

The present study aimed to assess the anti-leishmanial effects of curcumin nanoemulsion (CUR-NE) against Leishmania major (MRHO/IR/75/ER) in both in vitro and in vivo experiments. CUR-NE was successfully prepared via the spontaneous emulsification method. The in vitro effect of various concentrations of CUR-NE against L. major promastigotes was assessed using the flow cytometry method. In vivo experiments were carried out in BALB/c mice inoculated subcutaneously with 2 × 106 L. major promastigotes. Mice were treated with topical CUR-NE (2.5 mg/ml), intra-lesion injection of CUR-NE (2.5 mg/ml), topical CUR suspension (CUR-S, 2.5 mg/ml), topical NE without CUR (NE-no CUR), amphotericin B as the positive control group, and infected untreated mice as the negative control group. In vitro exposure of promastigotes to CUR-NE showed a dose-dependent anti-leishmanial effect, with a 67.52 ± 0.35% mortality rate at a concentration of 1250 µg/ml and an IC50 of 643.56 µg/ml. In vivo experiments showed that topical CUR-NE and CUR-S significantly decreased the mean lesion size in mice after four weeks from 4.73 ± 1.28 to 2.78 ± 1.28 mm and 4.45 ± 0.88 to 3.23 ± 0.59 mm, respectively (p = 0.001). Furthermore, CUR-NE significantly decreased the parasite load in treated mice compared with the negative control group (p = 0.001). Results from the current study demonstrated the promising activity of CUR-NE against L. major in both in vitro and in vivo experiments. Moreover, CUR-NE was more efficient than CUR-S in healing and reducing parasite burden in mouse models. Future studies should aim to identify molecular mechanisms as well as the pharmacologic and pharmacokinetic aspects of CUR-NE.

Designing Antitrypanosomal and Antileishmanial BODIPY Derivatives: A Computational and In Vitro Assessment.

Leishmaniasis and Human African trypanosomiasis pose significant public health threats in resource-limited regions, accentuated by the drawbacks of the current antiprotozoal treatments and the lack of approved vaccines. Considering the demand for novel therapeutic drugs, a series of BODIPY derivatives with several functionalizations at the meso, 2 and/or 6 positions of the core were synthesized and characterized. The in vitro activity against Trypanosoma brucei and Leishmania major parasites was carried out alongside a human healthy cell line (MRC-5) to establish selectivity indices (SIs). Notably, the meso-substituted BODIPY, with 1-dimethylaminonaphthalene (1b) and anthracene moiety (1c), were the most active against L. major, displaying IC50 = 4.84 and 5.41 µM, with a 16 and 18-fold selectivity over MRC-5 cells, respectively. In contrast, the mono-formylated analogues 2b and 2c exhibited the highest toxicity (IC50 = 2.84 and 6.17 µM, respectively) and selectivity (SI = 24 and 11, respectively) against T. brucei. Further insights on the activity of these compounds were gathered from molecular docking studies. The results suggest that these BODIPYs act as competitive inhibitors targeting the NADPH/NADP+ linkage site of the pteridine reductase (PR) enzyme. Additionally, these findings unveil a range of quasi-degenerate binding complexes formed between the PRs and the investigated BODIPY derivatives. These results suggest a potential correlation between the anti-parasitic activity and the presence of multiple configurations that block the same site of the enzyme.

In silico study and in vitro antileishmanial and antitrypanosomal evaluation of azopyrazoles and azopyrimidines.

Hydrazones 1-6, azo-pyrazoles 7-9 and azo-pyrimidines 10-15 are compounds that exhibit antibacterial activity. The mode of action and structures of these derivatives have been previously confirmed as antibacterial. In this investigation, biological screening and molecular docking studies were performed for derivatives 1-15, with compounds 2, 7, 8, 14 and 15 yielding the best energy scores (from -20.7986 to -10.5302 kcal/mol). Drug-likeness and in silico ADME prediction for the most potent derivatives, 2, 7, 8, 14 and 15, were predicted (from 84.46 to 96.85%). The latter compounds showed good recorded physicochemical properties and pharmacokinetics. Compound 8 demonstrated the strongest inhibition, which was similar to the positive control (eflornithine) against Trypanosoma brucei brucei (WT), with an EC50 of 25.12 and 22.52µM, respectively. Moreover, compound 14 exhibited the best activity against Leishmania mexicana promastigotes and Leishmania major promastigotes (EC50 =46.85; 40.78µM, respectively).

Identification and Validation of Compounds Targeting Leishmania major Leucyl-Aminopeptidase M17.

Leishmaniasis is a neglected tropical disease; there is currently no vaccine and treatment is reliant upon a handful of drugs suffering from multiple issues including toxicity and resistance. There is a critical need for development of new fit-for-purpose therapeutics, with reduced toxicity and targeting new mechanisms to overcome resistance. One enzyme meriting investigation as a potential drug target in Leishmania is M17 leucyl-aminopeptidase (LAP). Here, we aimed to chemically validate LAP as a drug target in L. major through identification of potent and selective inhibitors. Using RapidFire mass spectrometry, the compounds DDD00057570 and DDD00097924 were identified as selective inhibitors of recombinant Leishmania major LAP activity. Both compounds inhibited in vitro growth of L. major and L. donovani intracellular amastigotes, and overexpression of LmLAP in L. major led to reduced susceptibility to DDD00057570 and DDD00097924, suggesting that these compounds specifically target LmLAP. Thermal proteome profiling revealed that these inhibitors thermally stabilized two M17 LAPs, indicating that these compounds selectively bind to enzymes of this class. Additionally, the selectivity of the inhibitors to act on LmLAP and not against the human ortholog was demonstrated, despite the high sequence similarities LAPs of this family share. Collectively, these data confirm LmLAP as a promising therapeutic target for Leishmania spp. that can be selectively inhibited by drug-like small molecules.

Preparation and characterization of artemether-loaded niosomes in Leishmania major-induced cutaneous leishmaniasis.

Cutaneous leishmaniasis is the most prevalent form of leishmaniasis worldwide. Although various anti-leishmanial regimens have been considered, due to the lack of efficacy or occurrence of adverse reactions, design and development of novel topical delivery systems would be essential. This study aimed to prepare artemether (ART)-loaded niosomes and evaluate their anti-leishmanial effects against Leishmania major. ART-loaded niosomes were prepared through the thin-film hydration technique and characterized in terms of particle size, zeta potential, morphology, differential scanning calorimetry, drug loading, and drug release. Furthermore, anti-leishmanial effect of the preparation was assessed in vitro and in vivo. The prepared ART-loaded niosomes were spherical with an average diameter of about 100 and 300 nm with high encapsulation efficiencies of > 99%. The results of in vitro cytotoxicity revealed that ART-loaded niosomes had significantly higher anti-leishmanial activity, lower general toxicity, and higher selectivity index (SI). Half-maximal inhibitory concentration (IC50) values of ART, ART-loaded niosomes, and liposomal amphotericin B were 39.09, 15.12, and 20 µg/mL, respectively. Also, according to the in vivo study results, ART-loaded niosomes with an average size of 300 nm showed the highest anti-leishmanial effects in animal studies. ART-loaded niosomes would be promising topical drug delivery system for the management of cutaneous leishmaniasis.

Comparison of Intralesional Sodium Stibogluconate versus Intralesional Meglumine Antimoniate for the Treatment of Leishmania major Cutaneous Leishmaniasis.

Israel is endemic for Old-World cutaneous leishmaniasis. The most common species is Leishmania major. However, the available treatment options are limited. This study's objective was to compare the authors' experience with different antimony intralesional treatments of Leishmania major cutaneous leishmaniasis. A retrospective evaluation was undertaken for cases of Leishmania major cutaneous leishmaniasis treated by pentavalent antimony in a university-affiliated medical centre in Israel. The previous treatment of intralesional sodium stibogluconate (Pentostam®) was compared with the current treatment of meglumine antimoniate (Glucantime®). One hundred cases of cutaneous leishmaniasis were treated during the study period, of whom 33 were treated with intralesional sodium stibogluconate and 67 were treated with intralesional meglumine antimoniate. The patients were 78 males and 22 females, mean age 24 (range 10-67) and there was a total of 354 skin lesions. Within 3 months from treatment, 91% (30/33) of the intralesional sodium stibogluconate group and 88% (59/67) of the intralesional meglumine antimoniate group had complete healing of the cutaneous lesions after an average of 3 treatment cycles (non-statistically significant). In conclusion, the 2 different medications have the same efficacy and safety for treating cutaneous leishmaniasis. Pentavalent antimoniate intralesional infiltration treatment is safe, effective, and well tolerated with minimal side effects for Old-World cutaneous leishmaniasis.

Photodynamic therapy of cationic and anionic BSA-curcumin nanoparticles on amastigotes of Leishmania braziliensis and Leishmania major and Leishmania amazonensis.

Cutaneous leishmaniasis is a neglected disease prevalent in tropical countries, and conventional treatment can cause several serious side effects. Photodynamic therapy (PDT) can be considered a promising treatment alternative, as it is non-invasive therapy that has no side effects and uses accessible and low-cost substances, such as curcumin. This study evaluated the PDT response with cationic and anionic BSA nanoparticles encapsulated with curcumin in macrophages infected with L. braziliensis, L. major, and L. amazonensis. The nanoparticle system was characterized using a steady-state technique, scanning electron microscopy (SEM) study, and its biological activity was evaluated using macrophage cell lines infected with different Leishmania species. All spectroscopy measurements demonstrated that BSA curcumin (BSACur) has good photophysical properties, and confocal microscopy shows that macrophages and protozoa internalized the nanoparticles. The viability test demonstrated that at low concentrations, such as 0.1, 0.7, and 1.0 µmol. L-1, there was a decrease in cell viability after PDT application. Furthermore, a decrease in the number of parasites recovered was observed in the PDT groups. The results allowed us to conclude that curcumin loaded into BSA nanoparticles may have potential application in drug delivery systems for PDT protocols, demonstrating reduced cell viability at lower concentrations than free curcumin.

Investigating the Effect of Satureja khuzestanica Essential oil on MDR1 Gene Expression in Leishmania major.

BACKGROUND: Cutaneous leishmaniasis is among the neglected diseases in the world. Pentavalent antimonial compounds are considered the first-line treatment for this disease. However, using alternative natural products has received great attention due to the side effects of chemical drugs and drug resistance of the Leishmania parasite. The present study aims to investigate the effect of Satureja khuzestanica essential oil (SKEO) on MDR1 gene expression. METHODS: In this study, standard strains of Leishmania major promastigotes were exposed to 5, 10, 15, and 20 µg/ml of SKEO. MDR1 gene expression of parasites exposed to essential oil was evaluated using real-time PCR. GAPDH was employed as the housekeeping gene for internal control. RESULTS: Despite the increase, no statistically significant difference was observed in the relative expression of the MDR1 gene between the control group and the groups containing 5, 10, and 20 µg/ml of SKEO (P > 0.05). The relative expression of the MDR1 gene significantly increased in the group containing 15 µg/ml of essential oil compared to the control one (P < 0.05). CONCLUSION: This study showed that the use of essential oil of Satureja khuzestanica plant can have an increasing effect on the expression of MDR1 gene of Leishmania promastigotes, which is the best case if Satureja khuzestanica essential oil reduces the expression of MDR1 gene. So it seems that the use of essential oil of Satoria plant is effective in controlling Leishmania parasite, but its concentrations induce drug resistance. As a result, concentrations of essential oil should be used that have a controlling effect on the growth and proliferation of Leishmania parasite and also have the least effect on the induction of MDR1 gene expression.

Comparison of biosynthetic zinc oxide nanoparticle and glucantime cytotoxic effects on Leishmania major (MRHO/IR/75/ER).

Currently, zinc oxide (ZnO) particles are used in nanotechnology to destroy a wide range of microorganisms. Although pentavalent antimony compounds are used as antileishmanial drugs, they are associated with several limitations and side effects. Therefore, it is always desirable to try to find new and effective treatments. The aim of this research is to determine the antileishmanial effect of ZnO particles in comparison to the Antimoan Meglumine compound on promastigotes and amastigotes of Leishmania major (MRHO/IR/75/ER). After the extraction and purification of macrophages from the peritoneal cavity of C57BL/6 mice, L. major parasites were cultured in Roswell Park Memorial Institute-1640 culture medium containing fetal bovine serum (FBS) 10% and antibiotic. In this experimental study, the effect of different concentrations of nanoparticles was investigated using the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl-tetrazolium bromide (MTT) colorimetric method, in comparison to the glucantime on promastigotes, amastigotes and healthy macrophages in the culture medium. The amount of light absorption of the obtained color from the regeneration of tetrazolium salt to the product color of formazan by the parasite was measured by an enzyme-linked immunosorbent assay (ELISA) reader, and the IC50 value was calculated. IC50 after 24 h of incubation was calculated as IC50 = 358.6 µg/mL. The results showed, that the efficacy of ZnO nanoparticles was favorable and dose-dependent. The concentration of 500 µg/mL of ZnO nanoparticles induced 84.67% apoptosis after 72. Also, the toxicity of nanoparticles was less than the drug. Nanoparticles exert their cytotoxic effects by inducing apoptosis. They can be suitable candidates in the pharmaceutical industry in the future.

In vitro and In vivo Effects of Ethanolic Extract of Fumaria parviflora Lam. Embedded in Chitosan Nanoparticles Against Leishmania major.

INTRODUCTION: Fumaria has been traditionally used to treat skin damages due to anti-inflammatory properties. In the present study, we evaluated the effect of the ethanolic extract of Fumaria parviflora Lam. (F. parviflora) against Leishmania major (L. major) using chitosan biopolymer drug delivery system both In vitro and In vivo models. MATERIALS AND METHODS: The ethanolic extract of F. parviflora was analyzed by HPLC to determine its active ingredients content. The extract was then loaded on chitosan nanoparticles (CNPs). The parasite was treated with various concentrations of the ethanolic extract, CNPs and CNPs loaded with F. parviflora extract (CNPs@ F. parviflora). The size of lesions of treated mice were measured on a weekly basis. The parasite burden was evaluated 8 weeks after treatment. RESULTS: The HPLC analysis showed the presence of Fumaric acid at a high concentration. The percentage of the drug released from CNPs@ F. parviflora within 24 and 72 h were 65% and 90% respectively. The results showed that F. parviflora extract and CNPs@ F. parviflora caused 84% and 96% growth inhibition of L. major promastigotes as revealed by Neubauer chamber counting and MTT test respectively. The IC50 values of F. parviflora extract and CNPs@ F. parviflora were 450 and 68.4 µg/ml respectively. In amastigote assay, the best results showed in CNPs@ F. parviflora that only 2% of macrophages were infected with amastigotes. In vivo experiments for mice treated with F. parviflora and CNPs @ F. parviflora in comparison to control group showed a significant reduction (P < 0.05) in the mean diameter of the lesions (2.3 and 1.72 mm and 9.91 mm respectively). CONCLUSION: The ethanolic extract of F. parviflora both as standalone and loaded in CNPs showed promising inhibitory effects against L. major both upon In vitro and In vivo experimentation as well as therapeutic effects for wound healing in infected mice.

Tetrahydrobenzo[h]quinoline derivatives as a novel chemotype for dual antileishmanial-antimalarial activity graced with antitubercular activity: Design, synthesis and biological evaluation.

Derivatives with tetrahydrobenzo[h]quinoline chemotype were synthesized via one-pot reactions and evaluated for their antileishmanial, antimalarial and antitubercular activities. Based on a structure-guided approach, they were designed to possess antileishmanial activity through antifolate mechanism, via targeting Leishmania major pteridine reductase 1 (Lm-PTR1). The in vitro antipromastigote and antiamastigote activity are promising for all candidates and superior to the reference miltefosine, in a low or sub micromolar range of activity. Their antifolate mechanism was confirmed via the ability of folic and folinic acids to reverse the antileishmanial activity of these compounds, comparably to Lm-PTR1 inhibitor trimethoprim. Molecular dynamics simulations confirmed a stable and high potential binding of the most active candidates against leishmanial PTR1. For the antimalarial activity, most of the compounds exhibited promising antiplasmodial effect against P. berghei with suppression percentage of up to 97.78%. The most active compounds were further screened in vitro against the chloroquine resistant strain P. falciparum, (RKL9) and showed IC50 value range of 0.0198-0.096 µM, compared to IC50 value of 0.19420 µM for chloroquine sulphate. Molecular docking of the most active compounds against the wild-type and quadruple mutant pf DHFR-TS structures rationalized the in vitro antimalarial activity. Some candidates showed good antitubercular activity against sensitive Mycobacterium tuberculosis in a low micromolar range of MIC, compared to 0.875 µM of isoniazid. The top active ones were further tested against a multidrug-resistant strain (MDR) and extensively drug-resistant strain (XDR) of Mycobacterium tuberculosis. Interestingly, the in vitro cytotoxicity test of the best candidates displayed high selectivity indices emphasizing their safety on mammalian cells. Generally, this work introduces a fruitful matrix for new dual acting antileishmanial-antimalarial chemotype graced with antitubercular activity. This would help in tackling drug-resistance issues in treating some Neglected Tropical Diseases.

Antileishmanial effects and drugability characteristics of a heterocyclic copper complex: An in silico, in vitro and molecular study.

Leishmaniasis caused by the protozoan Leishmania presents a severe illness, principally in tropical and subtropical areas. Antileishmanial metal complexes, like Glucantime®ï¸ with proven activity, are routinely studied to probe their potency. We investigated the effects of a Cu (II) homoleptic complex coordinated by two dimethyl-bipyridine ligands against Leishmania major stages in silico and in vitro. The affinity of this heterocyclic Cu (II) complex (CuDMBP) towards a parasitic metacaspase was studied by molecular docking. Key pharmacokinetic and pharmacodynamic properties of the complex were predicted using three web-based tools. CuDMBP was tested for in vitro antileishmanial activities using MTT assay, model murine macrophages, flow cytometry, and quantitative real-time polymerase chain reaction (qPCR). Molecular docking confirmed the tendency between the target macromolecule and the complex. ADMET evaluations highlighted CuDMBP's key pharmacological features, including P-glycoprotein-associated GI absorption and lack of trans-BBB permeability. MTT showed significant inhibitory effects against promastigotes. CuDMBP significantly increased the level of cellular IL-12 expression (p < 0.05), while the upregulation observed in the expression of iNOS was considered not significant (p > 0.05). It decreased the expression of IL-10 significantly (p < 0.05). Findings demonstrated that CuDMBP deserves to be introduced as a leishmanicidal candidate provided further studies are carried out.

Discovery of novel Leishmania major trypanothione synthetase inhibitors by high-throughput screening.

Leishmaniasis is an infectious disease caused by obligate intracellular protozoa of the genus Leishmania with high infection and death rates in developing countries. New drugs with better pharmacological performance with regards to safety, efficacy, toxicity, and drug resistance than those/the ones currently used are urgently needed. Trypanothione synthetase (TryS) is an attractive target for the development of drugs against leishmaniasis because it is specific and essential to kinetoplastid parasites. In this study, Leishmaniamajor TryS was expressed and purified, and the kinetic parameters of purified TryS were determined. To identify novel inhibitors of LmTryS, a high-throughput screening (HTS) assay was developed and used to screen a library of 35,040 compounds. In the confirmatory assay, 42 compounds displayed half maximal inhibitory concentration (IC50) values < 50 µM and six of them corresponded to novel structures with IC50 ranging from 9 to 19 µM against LmTryS enzyme activity. Of the six inhibitors, TS001 showed the highest activity against growth of L. major promastigotes, L. donovani promastigotes, and Trypanosoma brucei brucei Lister 427 with IC50 values of 17, 26, and 31 µM, respectively. An in silico docking study using a homology model of LmTryS predicted the molecular interactions between LmTryS and the inhibitors.

Anti-leishmanial physalins-Phytochemical investigation, in vitro evaluation against clinical and MIL-resistant L. tropica strains and in silico studies.

Cutaneous leishmaniasis (CL) is a major health problem in over 98 countries of the world, including Pakistan. The current treatments are associated with a number of adverse effects and availability problem of drugs. Therefore, there is an urgent need of easily available and cost effective treatments of CL- in Pakistan. The bioassay-guided fractionation and purification of crude extract of Physalis minima has led to the isolation of a new aminophysalin B (1), and eight known physalins, physalin B (2), 5ß,6ß-epoxyphysalin B (3), 5α-ethoxy-6ß-hydroxy-5,6-dihydrophysalin B (4), physalin H (5), 5ß,6ß-epoxyphysalin C (6), and physalin G (7), K (8), and D (9). It is worth noting that compound 1 is the second member of aminophysalin series, whereas compound 6 was fully characterized for the first time. The structures of compounds 1-9 were elucidated by spectroscopic techniques Whereas, the structural assignments of compounds 1 and 8 were also supported by single-crystal X-ray diffraction studies. The anti-leishmanial activity of isolated physlains 1-9 was evaluated against Leishmania major and Leishmania tropica promastigotes. Compounds 2, 3, and 5-7 (IC50 = 9.59 ± 0.27-23.76 ± 1.10 µM) showed several-fold more potent activity against L. tropca than tested drug miltefosine (IC50 = 42.75 ± 1.03 µm) and pentamidine (IC50 = 27.20 ± 0.01 µM). Whereas compounds 2, 3 and 5 (IC50 = 3.04 ± 1.12-3.76 ± 0.85 µM) were found to be potent anti-leishmanial agents against L. major, several fold more active than tested standard miltefosine (IC50 = 25.55 ± 1.03 µM) and pentamidine (IC50 = 27.20 ± 0.015 µM). Compounds 4 (IC50 = 74.65 ± 0.81 µM) and 7 (IC50 = 39.44 ± 0.65 µM) also showed potent anti-leishmanial ativity against the miltefosine-unresponsive L. tropica strain (MIL resistant) (miltefosine IC50 = 169.55 ± 0.78 µM). Molecular docking and predictive binding studies indicated that these inhibitors may act via targeting important enzymes of various metabolic pathways of the parasites.

Multitarget, Selective Compound Design Yields Potent Inhibitors of a Kinetoplastid Pteridine Reductase 1.

The optimization of compounds with multiple targets is a difficult multidimensional problem in the drug discovery cycle. Here, we present a systematic, multidisciplinary approach to the development of selective antiparasitic compounds. Computational fragment-based design of novel pteridine derivatives along with iterations of crystallographic structure determination allowed for the derivation of a structure-activity relationship for multitarget inhibition. The approach yielded compounds showing apparent picomolar inhibition of T. brucei pteridine reductase 1 (PTR1), nanomolar inhibition of L. major PTR1, and selective submicromolar inhibition of parasite dihydrofolate reductase (DHFR) versus human DHFR. Moreover, by combining design for polypharmacology with a property-based on-parasite optimization, we found three compounds that exhibited micromolar EC50 values against T. brucei brucei while retaining their target inhibition. Our results provide a basis for the further development of pteridine-based compounds, and we expect our multitarget approach to be generally applicable to the design and optimization of anti-infective agents.

Broad-spectrum in vitro activity of macrophage infectivity potentiator inhibitors against Gram-negative bacteria and Leishmania major.

BACKGROUND: The macrophage infectivity potentiator (Mip) protein, which belongs to the immunophilin superfamily, is a peptidyl-prolyl cis/trans isomerase (PPIase) enzyme. Mip has been shown to be important for virulence in a wide range of pathogenic microorganisms. It has previously been demonstrated that small-molecule compounds designed to target Mip from the Gram-negative bacterium Burkholderia pseudomallei bind at the site of enzymatic activity of the protein, inhibiting the in vitro activity of Mip. OBJECTIVES: In this study, co-crystallography experiments with recombinant B. pseudomallei Mip (BpMip) protein and Mip inhibitors, biochemical analysis and computational modelling were used to predict the efficacy of lead compounds for broad-spectrum activity against other pathogens. METHODS: Binding activity of three lead compounds targeting BpMip was verified using surface plasmon resonance spectroscopy. The determination of crystal structures of BpMip in complex with these compounds, together with molecular modelling and in vitro assays, was used to determine whether the compounds have broad-spectrum antimicrobial activity against pathogens. RESULTS: Of the three lead small-molecule compounds, two were effective in inhibiting the PPIase activity of Mip proteins from Neisseria meningitidis, Klebsiella pneumoniae and Leishmania major. The compounds also reduced the intracellular burden of these pathogens using in vitro cell infection assays. CONCLUSIONS: These results indicate that Mip is a novel antivirulence target that can be inhibited using small-molecule compounds that prove to be promising broad-spectrum drug candidates in vitro. Further optimization of compounds is required for in vivo evaluation and future clinical applications.

Anti-leishmanial effects of resveratrol and resveratrol nanoemulsion on Leishmania major.

BACKGROUND: Leishmaniasis is a vector-borne disease that is endemic in the tropical and sub-tropical areas of the world. Low efficacy and high cytotoxicity of the current treatment regimens for leishmaniasis is one of the most important health problems. In this experimental study, anti-leishmanial effects of different concentrations of resveratrol and resveratrol nano-emulsion (RNE) were assessed. METHODS: RNE was prepared using the probe ultra-sonication method. The cytotoxicity was evaluated using the MTT technique on the L929 cell line. The anti-leishmanial activities on promastigotes of leishmania were assessed using vital staining and infected BALB/c mice were used to assess the in vivo anti-leishmanial effects. RESULTS: In vitro and in vivo assays revealed that all concentrations of resveratrol and RNE had valuable inhibitory effects against Leishmania major in comparison to the control group (P < 0.05). The half maximal inhibitory concentration (IC50) values were calculated as 16.23 and 35.71 µg/mL for resveratrol and RNE, respectively. Resveratrol and RNE showed no cytotoxicity against the L929 cell line. CONCLUSIONS: According to the potent in vitro and in vivo anti-leishmanial activity of RNE at low concentration against L. major, we suggest that it could be a promising anti-leishmanial therapeutic against L. major in the future.

Ellagitannins and simple phenolics from the halophytic plant Tamarix nilotica.

Three new [nilotinins M8‒M10 (1‒3)] and two known [tamarixinin A (4) and gemin D (5)] ellagitannins and seven simple phenolics [gallic acid (6), methyl gallate (7), 3,4-di-O-methylgallic acid (8), ellagic acid (9), 3-O-methylellagic acid (10), methyl ferulate 3-O-sulphate (11), and 7,4'-di-O-methylkaempferol (12)] were isolated from the halophytic plant Tamarix nilotica (Ehrenb.) Bunge (Tamaricaceae). Their structures were determined based on intensive spectroscopic studies and comparisons with reported data. Compounds 4, and 6-8 were evaluated for their cytotoxicity against lung adenocarcinoma cell line (A549) and anti-leishmanial activity against Leishmania major. Compounds 4, 6 and 7 showed promising cytotoxic properties against A549 (IC50 29 ± 2.3, 10.5 ± 0.7, and 20.7 ± 1.9 µg/mL), while compounds 4 and 7 showed higher growth-inhibitory effects against L. major promastigotes (IC50 40.5 ± 2.7 and 38.4 ± 2.5 µg/mL), as compared with the standards doxorubicin (IC50 0.42 µg/mL) and miltefosine (IC50 9.43 µg/mL), respectively.

Novel 3-chloro-6-nitro-1H-indazole derivatives as promising antileishmanial candidates: synthesis, biological activity, and molecular modelling studies.

An efficient pathway was disclosed for the synthesis of 3-chloro-6-nitro-1H-indazole derivatives by 1,3-dipolar cycloaddition on dipolarophile compounds 2 and 3. Faced the problem of separation of two regioisomers, a click chemistry method has allowed us to obtain regioisomers of triazole-1,4 with good yields from 82 to 90% were employed. Also, the antileishmanial biological potency of the compounds was achieved using an MTT assay that reported compound 13 as a promising growth inhibitor of Leishmania major. Molecular docking demonstrated highly stable binding with the Leishmania trypanothione reductase enzyme and produced a network of hydrophobic and hydrophilic interactions. Molecular dynamics simulations were performed for TryR-13 complex to understand its structural and intermolecular affinity stability in a biological environment. The studied complex remained in good equilibrium with a structure deviation of ∼1-3 Å. MM/GBSA binding free energies illustrated the high stability of TryR-13 complex. The studied compounds are promising leads for structural optimisation to enhance the antileishmanial activity.